Biology subjects

Cohen, E. A.

Publications and source records attributed to Cohen, E. A..

9 recordsLinked to original sources

bioRxiv2026

HIV-1 Vpu complements HLA-A downregulation by Nef and impairs TCR-mediated recognition of HIV-infected cells through distinct mechanisms

Many viruses, including HIV-1, target the human leukocyte antigen (HLA) class I molecules HLA-A, HLA-B, and HLA-C to evade immune surveillance. HIV-1 Nef selectively downregulates HLA-A and HLA-B, whereas Vpu variants from primary strains reduce cell-surface HLA-C expression. Whether HIV-1 also uses Vpu to target other HLA molecules has remained unclear. Here, we show that Vpu from primary strains reduces HLA-A, including the globally prevalent HLA-A2, on infected cells. Although Nef is the principal modulator of these molecules, Vpu acts independently of Nef and impairs TCR-mediated recognition of infected cells in co-culture systems. However, the reduction in TCR signaling is also dependent on the presence of BST2/tetherin, a restriction factor that retains nascent virions and targets them for degradation. HLA-A2-restricted, HIV-specific CTL responses were lowest when both Nef and Vpu were present. Vpu variants from chronic and transmitted/founder strains, but not from the prototypical recombinant strain NL4.3, were sufficient to reduce cell-surface HLA-A2, although activity varied widely among variants and appears to have arisen independently in different hosts. Coimmunoprecipitation and immunofluorescence analyses showed that Vpu interacts with HLA-A2 and redirects it to a perinuclear compartment which is probably the trans-Golgi network through a mechanism governed by the Vpu N-terminus/transmembrane domain. Overall, our findings reveal a previously unrecognized function of Vpu and show how HIV-1 uses Nef and Vpu to limit HLA-A-mediated viral antigen presentation and promote immune evasion.

immunology↗

bioRxiv2026

Impact of HIV-1 infection on the microglia transcriptome: induction of microRNA-23b leads to G3BP2-mediated stress granules assembly disruption

Microglia act as HIV reservoirs in the brain and contribute to the development of HIV-associated neurocognitive disorders despite antiretroviral therapy. We examined the transcriptome of primary microglia infected ex vivo with HIV-1. Analyses reveal a significant modulation of DNA damage/cell cycle-, cellular stress- and non-membrane bounded organelle assembly-related gene expression, and a broad effect on the microRNA profile. MicroRNA-23b is identified among upregulated microRNAs in HIV-1-infected microglia, and downregulated g3bp2 mRNA, validated as its target. G3BP2 is a core component of stress granules (SGs), which serve as antiviral and proinflammatory platforms. MicroRNA-23b mimics reduced both G3BP2 expression and SG assembly in stress-induced monocyte-derived microglial cells (MDMi), and few SGs aggregate in infected MDMi. Inhibition of MicroRNA-23b partially re-established SG formation and reduced HIV-1 replication. Thus, microRNA-23b induction contributes to HIV-1 strategies aimed at counteracting SG assembly in infected microglia, a host response that restricts HIV-1 replication and ensuing neuropathogenesis.

microbiology↗

bioRxiv2026

Comparative profiling of SARS-CoV-2 variant infections reveals diverse impacts on host cell RNA and RNA binding protein distribution and regulation

RNA viruses perturb cellular RNA metabolism to promote viral transcript expression and genome replication. However, the extent of host cell RNA reprogramming and the consequences on RNA binding protein (RBP) expression and localization remain poorly understood. Here, we utilized an integrated set of transcriptomics, proteomics, and high-content imaging approaches to compare the impact of different SARS-CoV-2 variants, including an early viral isolate and later emerging variants of concern (Alpha, Delta and Omicron) on the host cellular RNA and RBP landscapes. While all variants deeply remodelled the host transcriptome in a subcellular compartment-specific manner, we found that Omicron variants express lower levels of viral RNA products compared to earlier variants, while still driving efficient production of viral proteins. The surge in viral RNA expression correlated with altered subcellular RNA biotype distribution, cytoplasmic mRNA exclusion, the selective disruption of the localization of polyA-tail interacting proteins, and the nuclear accumulation of non-coding transcripts such as RN7SL family members and MALAT-1. Unexpectedly, we found many RBPs that exhibiting altered subcellular localization following abiotic innate immune stimulation with poly(I:C) but remained unaltered in cells infected with SARS-CoV-2 variants, revealing a capacity for these viruses to limit the overall impact on the RBP subcellular distribution. Our results further indicate that Omicron infection sustains higher expression levels of proteins components of the signal recognition particle and translation factors, which may enable a more effective production of immune response and viral components. Altogether, this work highlights the deep impact of SARS-CoV-2 infection on host cell RNA regulatory landscape and provides new insights to explain the differential impact of Omicron compared to early SARS-CoV-2 variants. HighlightsO_LISARS-CoV-2 variants drive varying degrees of viral RNA expression in infected Calu-3 cells C_LIO_LIInfection reshapes the host cell RNA landscape in a cell-compartment specific manner C_LIO_LIRN7SL RNA exhibits a strong increase in relative abundance and accumulates in the nucleus of infected cells C_LIO_LISARS-CoV-2 infections impact the localization of polyA-tail binding proteins while blocking the nuclear accumulation of immune response-sensitive RBPs C_LIO_LIOmicron infections are less disruptive to cellular function and sustain the production of translation factors and components of the SRP complex C_LI

molecular biology↗

bioRxiv2025

Humanized Bone Marrow-Liver-Thymus Mice for Studying HIV-1 Persistence in Liver and Lung CD4+ T and Myeloid Cell Subsets during Antiretroviral Therapy

BackgroundWhile the role of CD4+ T-cells in HIV-1 reservoir persistence during antiretroviral therapy (ART) is well-established, studies on tissue-resident macrophages (M{Phi}) in people with HIV-1 (PWH) are restricted by difficulties in accessing deep tissue samples. Investigations in myeloid-only humanized mouse models demonstrated the contribution of M{Phi} to viral rebound upon ART interruption. Two distinct M{Phi} subsets exist in mice and humans: one of embryonic origin and self-renewal capacity generating long-lived tissue-resident M{Phi} (LL-TRM), and another one of short-lived M{Phi} (SL-M{Phi}), constantly replenished by bone marrow monocytes. The relative contribution of LL-TRM versus SL-M{Phi} to tissue HIV-1 reservoir persistence during ART remains understudied. Here, we used a humanized BM-liver-thymus (hu-BLT) mouse model to quantify integrative HIV-1 infection in liver/lung M{Phi} versus CD4+ T-cells before/after ART and document their expression of LL-TRM/SL-M{Phi} markers. ResultsLung/liver immune cells were extracted from ART-naive (ART-) and ART-treated (ART+) HIV-infected (HIV+) hu-BLT mice, as well as HIV-uninfected mice (HIV-). M{Phi} were identified as cells expressing the myeloid markers CD33/HLA-DR and/or CD68 and flow-cytometry sorted based on their differential expression of CD14 and/or CCR2. Matched CD3+CD4+ T-cells were sorted in parallel and used as controls. HIV-DNA integration was measured by nested real-time PCR. In contrast to CD4+ T-cells that carried the highest levels of proviral HIV-DNA before and after ART, integrative infection in liver/lung M{Phi} was detected before ART, but was drastically reduced in HIV+ART+ hu-BLT mice, regardless of CD14 or CCR2 expression on M{Phi}. Markers of LL-TRM (CD163/CX3CR1/Ki67/c-Kit) were expressed on a small fraction of liver but not lung M{Phi}, indicative of a deficient LL-TRM development in this hu-BLT model. ConclusionsTogether our results demonstrate that lung/liver M{Phi} in hu-BLT mice support integrative HIV-1 infection in vivo, but their contribution to viral reservoir persistence during ART is minor when compared to CD4+ T-cells. This is consistent with the deficient development of LL-TRM we observed in hu-BLT mice. However, HIV-1 permissive M{Phi} present in this model likely contribute to viral rebound upon ART interruption. Therefore, HIV-1 cure interventions that are tested in such preclinical models should consider targeting HIV-1 replication in both M{Phi} and CD4+ T-cells.

immunology↗

bioRxiv2022

Generation of Functional Human T Cell Development in NOD/SCID/IL2rγnull Humanized Mice Without Using Fetal Tissue: Application as a Model of HIV Infection and Persistence

Generating humanized mice with fully functional T cells currently relies on co-implantation of hematopoietic stem cells from fetal liver and autologous fetal thymic tissue (BLT mouse). However, access to such tissues has ethical and logistical challenges. Herein, we show that NOD/SCID/IL2r{gamma}null mice humanized with cord blood-derived CD34+ cells and implanted in quadriceps with pediatric thymic tissues excised during cardiac surgeries (CCST mice) are an alternative to BLT mice. Our data revealed a strong immune reconstitution in CCST mice, with T cells originating from CD34+ progenitor cells, proliferating efficiently in response to mitogenic stimulation ex vivo and capable of rejecting allogeneic human leukemic cells in vivo. Despite having less T cells than BLT mice, CCST mice were equally susceptible to mucosal or intraperitoneal HIV infection. Importantly, HIV-specific T cell responses were significantly higher in CCST-mice (median: 10.4% vs. 0.7%; p<0.0001 for CD8+cells and 3.9% vs. 0.7%; p<0.01 for CD4+ cells). As well, antiretroviral therapy (ART) robustly suppressed viremia and reduced the frequencies of cells carrying integrated HIV DNA by up to 2 logs in various CCST mouse tissues. As in BLT mice, we observed a complete viral rebound in 67% of the animals by 2-4 weeks following ART interruption, suggesting the presence of HIV reservoirs. In conclusion, CCST mice represent an ethical and practical alternative to BLT mice, broadening the feasibility of utilizing humanized mice for research on HIV and other human diseases. One Sentence SummaryWe herein report a new humanized mouse model implanted with human cord blood hematopoietic stem cells and allogenic pediatric thymic tissue that develops a functional T cell compartment and supports efficient HIV infection and persistence during antiretroviral therapy.

immunology↗

bioRxiv2021

Implications of Spike-glycoprotein processing at S1/S2 by Furin, at S2' by Furin and/or TMPRSS2 and shedding of ACE2: cell-to-cell fusion, cell entry and infectivity of SARS-CoV-2

Disclaimer StatementThe author has withdrawn this manuscript due to a duplicate posting of manuscript number 423106. Therefore, the author does not wish this work to be cited as reference for the project. If you have any questions, please contact the corresponding author (Nabil G. Seidah at seidahn@ircm.qc.ca.

biochemistry↗

bioRxiv2021

HIV-1 Vpu promotes phagocytosis of infected CD4+ T cells by macrophages through downregulation of CD47

Human immunodeficiency virus (HIV) remodels the cell surface of infected cells to facilitate viral dissemination and promote immune evasion. The membrane-associated Vpu accessory protein encoded by HIV-1 plays a key role in this process by altering cell surface levels of multiple host proteins. Using an unbiased quantitative plasma membrane profiling approach, we previously identified CD47 as a putative host target downregulated by Vpu. CD47 is a ubiquitously-expressed cell surface protein that interacts with the myeloid cell inhibitory receptor SIRP to deliver a "dont-eat-me" signal, thus protecting cells from phagocytosis. In this study, we investigate whether CD47 modulation by HIV-1 Vpu might promote the susceptibility of macrophages to viral infection via phagocytosis of infected CD4+ T cells. Indeed, we find that Vpu downregulates CD47 expression on infected CD4+ T cells leading to an enhanced capture and phagocytosis by macrophages. Interestingly, it is through this process that a CCR5-tropic transmitted/founder (T/F) virus, which otherwise poorly infects macrophages in its cell-free form, becomes infectious in macrophages. Importantly, we show that HIV-1-infected cells expressing a Vpu-resistant CD47 mutant are less prone to infect macrophages through phagocytosis. Mechanistically, Vpu forms a physical complex with CD47 through its transmembrane domain and targets the latter for lysosomal degradation. These results reveal a novel role of Vpu in modulating macrophage infection, which has important implications for HIV-1 transmission in early stages of infection and the establishment of viral reservoir. IMPORTANCEMacrophages play critical roles in HIV transmission, viral spread early in infection, and as a reservoir of virus. Selective capture and engulfment of HIV-1 infected T cells was shown to drive efficient macrophage infection suggesting that this mechanism represents an important mode of infection notably for weakly macrophage-tropic T/F viruses. In this study, we provide insight into the signals that regulate this process. We show that the HIV-1 accessory protein Vpu downregulates cell surface levels of CD47, a host protein that interacts with the inhibitory receptor SIRP to deliver a "dont-eat-me" signal to macrophages. This allows for enhanced capture and phagocytosis of infected T cells by macrophages, ultimately leading to their productive infection even with T/F virus. These findings provide new insights into the mechanisms governing the intercellular transmission of HIV-1 to macrophages with implications for the establishment of the macrophage reservoir and early HIV-1 dissemination in vivo.

microbiology↗

bioRxiv2020

Furin cleaves SARS-CoV-2 spike-glycoprotein at S1/S2 and S2'for viral fusion/entry: indirect role for TMPRSS2

The spike (S)-protein of SARS-CoV-2 binds ACE2 and requires proteolytic "priming" at PRRAR685{downarrow} into S1 and S2 (cleavage at S1/S2), and "fusion-activation" at a S2 site for viral entry. In vitro, Furin cleaved peptides mimicking the S1/S2 cleavage site more efficiently than at the putative S2, whereas TMPRSS2 inefficiently cleaved both sites. In HeLa cells Furin-like enzymes mainly cleaved at S1/S2 during intracellular protein trafficking, and S2 processing by Furin at KPSKR815{downarrow} was strongly enhanced by ACE2, but not for the optimized S2 KRRKR815{downarrow} mutant (S2), whereas individual/double KR815AA mutants were retained in the endoplasmic reticulum. Pharmacological Furin-inhibitors (Boston Pharmaceuticals, BOS-inhibitors) effectively blocked endogenous S-protein processing in HeLa cells. Furthermore, we show using pseudotyped viruses that while entry by a "pH-dependent" endocytosis pathway in HEK293 cells did not require Furin processing at S1/S2, a "pH-independent" viral entry in lung-derived Calu-3 cells was sensitive to inhibitors of Furin (BOS) and TMPRSS2 (Camostat). Consistently, these inhibitors potently reduce infectious viral titer and cytopathic effects, an outcome enhanced when both compounds were combined. Quantitative analyses of cell-to-cell fusion and spike processing revealed the key importance of the Furin sites for syncytia formation. Our assays showed that TMPRSS2 enhances fusion and proteolysis at S2 in the absence of cleavage at S1/S2, an effect that is linked to ACE2 shedding by TMPRSS2. Overall, our results indicate that Furin and TMPRSS2 play synergistic roles in generating fusion-competent S-protein, and in promoting viral entry, supporting the combination of Furin and TMPRSS2 inhibitors as potent antivirals against SARS-CoV-2. IMPORTANCESARS-CoV-2 is the etiological agent of COVID-19 that resulted in >5 million deaths. The spike protein (S) of the virus directs infection of the lungs and other tissues by binding the angiotensin-converting enzyme 2 (ACE2) receptor. For effective infection, the S-protein is cleaved at two sites: S1/S2 and S2. Cleavage at S1/S2, induces a conformational change favoring the recognition of ACE2. The S2 cleavage is critical for cell-to-cell fusion and virus entry into host cells. Our study contributes to a better understanding of the dynamics of interaction between Furin and TMPRSS2 during SARS-CoV-2 entry and suggests that the combination of a non-toxic Furin inhibitor with a TMPRSS2 inhibitor could significantly reduce viral entry in lung cells, as evidenced by an average synergistic [~]95% reduction of viral infection. This represents a powerful novel antiviral approach to reduce viral spread in individuals infected by SARS-CoV-2 or future related coronaviruses.

biochemistry↗

bioRxiv2019

ILT7 activation and plasmacytoid dendritic cell response are governed by BST2 determinants that are structurally-distinct

The interaction of the human pDC receptor ILT7 with its ligand, BST2, significantly regulates pDCs TLR-induced innate immune responses. This interaction has critical biological consequences, yet, its structural requirements are not fully characterized. The BST2 ectodomain can be divided in structurally and functionally distinct regions; while the coiled-coil region contains a newly-defined ILT7 binding surface, the N-terminal region appears to negatively modulate ILT7 activation. A stable BST2 homodimer binds to ILT7 but post-binding events associated to the unique BST2 coiled-coil plasticity are required to trigger receptor signaling. Hence, BST2 with an unstable or with a rigid coiled-coil fails to activate ILT7, whereas mutations in the N-terminal region enhance activation. Importantly, the biological relevance of these newly defined domains of BST2 is underscored by the identification of mutations with opposed potential to activate ILT7, which are selected under pathological malignant conditions.

immunology↗